Polymorphisms in CD1a and susceptibility to mTb
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Polymorphisms in CD1a and susceptibility to mTb
Emily Ford5/29/2012
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Acknowledgments:
• Thomas Hawn Chetan SeshadriMeera ShenoyGlenna PetersonRick Wells
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NA 1/34McMichael AJ, JR Pilch, G Galfre, DY Mason, JW Fabre, C Milstein. 1979. Eur J Immunol 9 (3): 205-210. A human thymocyte antigen defined by a hybrid myeloma monoclonal antibody.
1984 – Nobel Prize - Niels K. Jerne, Georges J.F. Kohler, Cesar Milstein
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“The dissection of the immune
response by the hybridoma technique”
Milstein C. Nobel prize lecture. 1984.
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Cluster of Differentiation (CD)
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Cluster analysisof the original 139 human antibody/antigen complexes
Bernard, A and L Boumsell (1984). Human Immunology 11: 1-10. IUIS-WHO Nomenclature Subcommittee (1984). Bull WHO 62 (5): 809-811.
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CD1
• Sequence lies on chromosome 1• Significant sequence homology with MHC-I• Associates with beta 2-microglobulin • Expressed on antigen presenting cells (APCs)
Porcelli S, MB Brenner et al. (1989). Nature 341 (6241): 447-450.Brigl M and MB Brenner. (2004). Annu Rev Immunol 22: 817-890.
CD1D CD1A CD1C CD1B CD1E
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MOLT-4(CD1)
IDP2 IDP2
IDP2
IDP2
IDP2
Brenner MB, J McLean et al. (1987.) Nature 325: 389-394
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CytotoxicCD4-8- T cells
CD1-transfected cells
CD1cPorcelli S, MB Brenner et al. (1989). Nature 341 (6241): 447-450.
IDP2
BK6
CD1-expressing cells can be recognized and lysed by specific CTLs
CD1a
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CTLs will respond to MTb in the presence of CD1
Porcelli, S, CT Morita, MB Brenner. (1992). Nature 360: 593-597
APC
DN1
DN1
DN1
DN1
?
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CD1 presents lipid antigens
Beckman EM, SA Porcelli et al. 1994. Nature 372: 691-4
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CD8+ CD1-responsive T cells can cause intracellular lysis of mycobacteria
Stenger S, RJ Mazzaccaro et al. 1997. Science 276: 1684-1687.
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CD1 family structure
Van Rhijn I, DM Zajonc et al. (2005). Curr Opin Immunol 17(3): 22-29.
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Sugita, M, DB Moody et al. 1998. Clinical Immunology and Immunopathology 87 (1): 8-14.
Overview and homology w/ known MHC presenting processes
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Tb: How is it so good at getting around our immune system?
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Coding region
Untranslated region
Are SNPs in CD1a Associated with Expression?
• Experimental Method– Collect blood from healthy individuals in Seattle (N=21)– Genotype for CD1a polymorphisms– Generate dendritic cells and analyze surface expression of CD1a
rs31
8102
9
rs85
8998
rs41
1089
rs22
6971
4
rs44
0419
rs38
9293
rs12
3071
6
Chromosome 1158,223,927 158,228,059
Peripheral BloodMononuclear Cells
CD1- Monocytes
CD1+
dendritic cellsGM-CSFIL-4
C Seshadri, not yet published
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Distribution of CD1a expression in Seattle cohort
Donor 3
Donor 4
Donor 5
CD1a CD1c
C Seshadri, not yet published
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CD1A Expression Stratified by SNP
Genotypic Recessive
p = 0.02 p = 0.006
Genotypic Recessive
p = 0.764 p = 0.472
Genotypic Test = Kruskal-Wallis rank testRecessive Test = Wilcoxon rank sums testC Seshadri, not yet published
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Is CD1a-deficiency Associated with Tuberculosis?
• Combined Cohort Analysis– 762 Cord Blood Controls– 385 Pulmonary TB– 307 Meningeal TB
SNP Cohort Genotypic Analysis Recessive Analysis
TT CT CC P value TT+CT CC OR P value
rs858998 control 372(49%) 327(43%) 59(8%) 699(92%) 59(8%)
T/C TB all 337(49%) 271(39%) 82(12%) 0.023 608(88%) 82(12%) 1.60 0.009
pTB 188(49%) 149(39%) 47(12%) 0.037 337(88%) 47(12%) 1.65 0.014
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Hypothesis: differences in the promoter region of CD1a may affect its expression
Promoter Intron Exon
Chromosome 1 – 5000kb
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PGL4
1. Make plasmids containing CD1a variants
CD1a “HI”
CD1a “LO”
1kb
1kb
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2. Try to get it into a cell
Lipofection Electroporation
cellcell
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Results
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The end!
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• If there’s time – mysterious CD1d
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• How did they move into dendritic cells?• They first had to make their own CD1 expressing cells,
then technology developed
• Ralph Steinman – PNAS from 1980s? – Dendritic cells were very hard to study! (rare, difficult to isolate) – Wesley Van Voorhees was his phd student
– GM-CSF IL-4 method to make dendritic cells (legend of figure 1) Nature paper
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Human thymocytesNA 1/34
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NA 1/34
HTA1/CD1
Human thymocytes
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Other T-cell types also recognize CD1
• Reminder of T cell subsets
CD4-8-
CD4+
CD8+